Quantitative assay for mutation in diploid human lymphoblasts using microtiter plates

Furth, Emma E.; Thilly, William G.; Penman, Bruce W.; Liber, Howard L.; Rand, William

doi:10.1016/0003-2697(81)90103-2

Cited by 248 publications

(105 citation statements)

References 16 publications

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“…Plates were scored for colony growth using an inverted microscope. The cloning efficiencies and mutant frequencies were estimated as described (Furth et al, 1981).…”

Section: Methodsmentioning

confidence: 99%

Radiation-induced transgenerational alterations in genome stability and DNA damage

et al. 2006

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Mutation induction in directly exposed cells is currently regarded as the main component of the genetic risk of ionizing radiation for humans. However, recent data on the transgenerational increases in mutation rates in the offspring of irradiated parents indicate that the genetic risk could be greater than predicted previously. Here, we have analysed transgenerational changes in mutation rates and DNA damage in the germline and somatic tissues of non-exposed first-generation offspring of irradiated inbred male CBA/Ca and BALB/c mice. Mutation rates at an expanded simple tandem repeat DNA locus and a proteincoding gene (hprt) were significantly elevated in both the germline (sperm) and somatic tissues of all the offspring of irradiated males. The transgenerational changes in mutation rates were attributed to the presence of a persistent subset of endogenous DNA lesions (double-and singlestrand breaks), measured by the phosphorylated form of histone H2AX (c-H2AX) and alkaline Comet assays. Such remarkable transgenerational destabilization of the F 1 genome may have important implications for cancer aetiology and genetic risk estimates. Our data also provide important clues on the still unknown mechanisms of radiation-induced genomic instability.

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“…Plates were scored for colony growth using an inverted microscope. The cloning efficiencies and mutant frequencies were estimated as described (Furth et al, 1981).…”

Section: Methodsmentioning

confidence: 99%

Radiation-induced transgenerational alterations in genome stability and DNA damage

et al. 2006

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“…TK-6 cells (human B lymphoblast; TK, thymidine kinase) were grown in RPMI medium 1640 supplemented with 10% horse serum and antibiotics. Details for assessing survival and mutagenicity at the TK locus have been described (22,23). TK-6 cells were treated with 0, 100, 120, and 140 M of 4-OHE 2 for 2 h in the absence or presence of 1 mM cysteine, 5 mM TPH, or 1 mM ascorbate.…”

Section: Methodsmentioning

confidence: 99%

Semiquinone radical intermediate in catecholic estrogen-mediated cytotoxicity and mutagenesis: Chemoprevention strategies with antioxidants

Samuni

Chuang

Krishna

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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Modulation of the cytotoxicity and mutagenicity of 4-hydroxyestradiol (4-OHE2), an oxidative metabolite of estrogen, by antioxidants was assessed in human MCF7 cells and TK-6 lymphoblast cells. The cytotoxicity of the catecholic estrogens was potentiated by depletion of intracellular glutathione and was independent of oxygen concentration. Agents such as the nitroxide Tempol can facilitate the oxidation of the semiquinone to the Q and enhanced 4-OHE2 cytotoxicity. Conversely, reducing agents such as ascorbate, cysteine, and 1,4-dihydroxytetramethylpiperidine (THP) protected against cytotoxicity and decreased mutation induction, presumably by reducing the semiquinone to the hydroquinone. Our results support the proposition that oxidation of the semiquinone to the corresponding Q is crucial in eliciting the deleterious effects of catecholic estrogens. Furthermore, because the deleterious effects of 4-OHE 2 were abrogated by dietary and synthetic antioxidants, our results would support the chemopreventive use of diets rich in reducing substances (vitamins and added synthetic antioxidants) as a means of decreasing the risks associated with estrogen exposure and developing of breast cancer.

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“…CE was calculated using Poisson statistics as described previously (Furth et al, 1981), where CE ¼ [Àln (fraction of negative wells)]/(number of cells seeded per well). The CE for each cell line presented in Table 1 is expressed as the mean for all replicate cultures of that line.…”

Section: Cell Linesmentioning

confidence: 99%

“…Mutation frequency (MF) and mutation rate (MR) calculations were performed essentially as described . Briefly, mutation rates were calculated using Poisson corrected counts (Furth et al, 1981) and the tables of Capizzi and Jameson (Capizzi and Jameson, 1973). For cell lines lacking mutations, mutation frequencies and rates were calculated assuming that one hypothetical mutant had been isolated and expressed as less than that value.…”

Section: Cell Linesmentioning

confidence: 99%

A subgroup of microsatellite stable colorectal cancers has elevated mutation rates and different responses to alkylating and oxidising agents

Parker

Leonard

et al. 2004

Br J Cancer

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An early step in the carcinogenesis of hereditary non-polyposis colorectal cancer (HNPCC) and some sporadic colorectal cancers (CRCs) is the acquisition of a 'mutator phenotype' resulting from defects in DNA mismatch repair (MMR) genes, which normally maintain genomic stability. This mutator phenotype causes an approximately 100 -1000-fold increase in base substitutions and small insertion/deletion mutations thereby driving carcinogenesis. It also causes genome-wide microsatellite instability (MSI) due to the inability to repair mutations within these small, hard to replicate, repetitive DNA elements. In contrast, less is known about the role of mutator phenotypes in microsatellite stable (MSS) CRC. In this report, we have measured the mutation rates in 11 MSS CRC cell lines to obtain an estimate of the prevalence of mutator phenotypes in MSS carcinogenesis. Of the 11 cell lines, three of them (27%) possess spontaneous hypoxanthine phosphoribosyltransferase mutation rates approximately 10 -100-fold above background. When challenged with alkylating and oxidising agents, the degree of survival and apoptotic responses are different, indicating that these cell lines may represent more than one mutator phenotype. These data demonstrate that a significant portion of MSS CRC cell lines has increased mutation rates and that this may play a role in MSS CRC carcinogenesis.

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Quantitative assay for mutation in diploid human lymphoblasts using microtiter plates

Cited by 248 publications

References 16 publications

Radiation-induced transgenerational alterations in genome stability and DNA damage

Radiation-induced transgenerational alterations in genome stability and DNA damage

Semiquinone radical intermediate in catecholic estrogen-mediated cytotoxicity and mutagenesis: Chemoprevention strategies with antioxidants

A subgroup of microsatellite stable colorectal cancers has elevated mutation rates and different responses to alkylating and oxidising agents

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